Theasaponin E1 destroys the salt tolerance of yeasts

Cells of Zygosaccharomyces rouxii in a medium containing a high concentration of NaCl were killed during incubation for 2-4 h with a low concentration of a mixture of saponins from tea seeds (TSS). The higher the concentration of NaCl in the medium, the higher the inhibitory effect of TSS on the growth of the yeast. The above inhibitory effect of TSS on the growth of the yeast was not observed when cells were incubated in hypertonic media composed of nonionic substances such as sugars. The ATPase activity of plasma membrane preparations from the yeast cells was slightly affected by the addition of TSS. It is shown that TSS facilitates leakage of glycerol from the yeast cells under NaCl-hypertonic conditions. The major inhibitor in the mixture of saponins was isolated and identified as theasaponin E1. Its isomer, theasaponin E2, did not have any effect on the salt tolerance of Z. rouxii or Saccharomyces cerevisiae.     

Characterization of plasma membrane proton-ATPase from salt-tolerant yeast Zygosaccharomyces rouxii cells

Plasma membrane was isolated from the salt-tolerant yeast Zygosaccharomyces rouxii and from Saccharomyces cerevisiae. The ATPase in the plasma membrane of Z. rouxii cells was a typical proton-ATPase as judged by testing with various ATPase inhibitors. There were slight differences in the pH optima of activities and in the sensitivity to sodium chloride (NaCl) and potassium chloride (KCl) of the ATPase from Z. rouxii and S. cerevisiae. The specific ATPase activity from Z. rouxii was higher in cells grown in a medium containing 2 _M-NaCl than in those not containing NaCl. No in vivo activation by incubation with glucose was observed in Z. rouxii cells and the specific ATPase activity was independent of the growth phase, unlike S. cerevisiae cells.     

Characterization of plasma membrane H+-ATPase from salt-tolerant yeast Candida versatilis

Plasma membrane was isolated from the salt-tolerant yeast Candida versatilis and the ATPase in plasma membrane was characterized. The ATPase was a typical H⁺-ATPase with similar properties to the Saccharomyces cerevisiae and Zygosaccharomyces rouxii enzymes. It was reacted with antibody (IgG) raised against S. cerevisiae plasma membrane H⁺-ATPase. The ATPase activity was not changed by adding NaCl and KCl to the assay solutions, but was increased by NH, especially by ammonium sulfate. In vivo stimulation of ATPase activity was observed by the addition of NaCl into the culture medium, as observed in Z. rouxii. No in vivo activation of H⁺-ATPase by glucose metabolism was observed in C. versatilis cells and the activity was independent of the growth phase, like Z. rouxii and unlike S. cerevisiae cells.     

The influence of precultivation parameters on the catabolism of branched-chain amino acids by Staphylococcus xylosus and Staphylococcus carnosus

The influence of precultivation parameters on the ability of Staphylococcus xylosus and Staphylococcus carnosus to convert branched-chain amino acids—leucine, isoleucine and valine—into volatile flavour compounds was investigated using resting cells in a defined reaction medium. The studied precultivation parameters were: growth phase, temperature, NaCl concentration and the concentration of leucine, isoleucine and valine (only for S. xylosus). Flavour compounds were sampled by automatic static headspace collection and separated/quantified using gas chromatography/flame ionization detection (GC/FID).Main catabolic products from degradation of leucine, isoleucine and valine were the flavour intensive branched-chain acids: 2- and 3-methylbutanoic and 2-methylpropanoic acids. The precultivation parameters altered the production of the branched-chain acids significantly, but to various degrees for S. xylosus and S. carnosus.Production of branched-chain acids by S. carnosus was only influenced slightly by the growth phase and not by changing the NaCl concentration between 4.0% and 10.0% (w/w). Lowering the temperature from 28°C to 18°C significantly decreased S. carnosus’ generation of branched-chain acids. In contrast, S. xylosus was significantly influenced by all precultivation parameters, in particular by the growth phase. Cells taken from growing cultures had a much higher production of branched-chain acids compared to cells taken from stationary cultures. Addition of leucine and valine to the precultivation medium enhanced the production of branched-chain acids whereas addition of isoleucine had the opposite effect.     

Development of defined mixed-culture fungal fermentation starter granulate for controlled production of rice wine

As a first step in the development of defined fungal starter granules for controlled winemaking from purple glutinous rice, the interaction of moulds and yeasts isolated from Vietnamese rice wine starters and the effect of some representative oriental herbs on the growth of moulds and yeasts were examined. Amylomyces rouxii and Saccharomyces cerevisae were shown to be compatible in mixed cultures, and the herbs “Tieu Hoi” (Fennel: Foeniculum vulgare Miller) and “Dinh Huong” (Clove: Syzygium aromaticum L.) which are used as supplementary ingredients by some local starter producers, were observed to stimulate the mould and yeast growth. Based on traditional starter manufacturing methods and modified on the basis of optimization experiments, a laboratory-scale manufacturing process for defined mixed-culture starter granules was established. In accordance with the national standard method, the wine produced with new experimental starter granules was found to have superior flavour and overall acceptability, compared with local commercial rice wines.

Industrial relevance

One of the major problems faced by commercial brewers of rice wine in Vietnam, is the variable quality and performance of the traditional starter tablets that are commonly used. The relevance of the present paper is that a stable, granulated starter has been developed, containing a defined mixture of mould and yeast cultures. This has proven to be shelf stable for more than 3 months, producing a very well accepted quality of wine.     

Effects of process parameters on growth and metabolism of<Emphasis Type="Italic"> Lactobacillus sanfranciscensis</Emphasis> and<Emphasis Type="Italic"> Candida humilis</Emphasis> during rye sourdough fermentation

The effects of temperature, pH, inoculum level, and NaCl on the growth and metabolism of Lactobacillus sanfranciscensis and Candida humilis in rye sourdough were determined. The temperature optima for growth of C. humilis and L. sanfranciscensis were 28 and 32 °C, respectively. Yeast growth was inhibited at 35 °C. The pH did not affect yeast growth in the range 3.5–5.5, whereas growth of L. sanfranciscensis was inhibited at pH 4.0. A NaCl concentration of 4% (flour base) inhibited growth of L. sanfranciscensis but not C. humilis. The effects of the process parameters on the formation of lactate, acetate, ethanol, and CO₂ by the organisms were generally in agreement with their effects on growth. However, decreased formation of acetate by L. sanfranciscensis was observed at 35 °C although lactate and ethanol formation were not affected. In conclusion, the study provides a rationale for the stable persistence of L. sanfranciscensis and C. humilis in traditional sourdoughs and will facilitate the optimisation of sourdough fermentations in traditional and new applications.     

On the selection of relevant environmental factors to predict microbial dynamics in solidified media

Several studies have shown that food structure causes slower growth rates and narrower growth boundaries of bacteria compared to laboratory media. In predictive microbiology, both a_w or corresponding solute concentration (mainly NaCl) have been used as a growth influencing factor for kinetic models or growth/no growth interface models. The majority of these models have been based on data generated in liquid broth media with NaCl as the predominant a_w influencing solute. However, in complex food systems, other a_w influencing components might be present, next to NaCl. In this study, the growth rate of Salmonella typhimurium was studied in the growth region and the growth/no growth response was tested in Tryptic Soy Broth at 20 °C at varying gelatin concentration (0, 10, 50 g L⁻¹ gelatin), pH (3.25–5.5) and water activity (a_w) (0.929–0.996). From the viewpoint of water activity, the results suggest that NaCl is the main a_w affecting compound. However, gelatin seemed to have an effect on medium a_w too. Moreover, there is also an interaction effect between NaCl and gelatin. From the microbial viewpoint, the results confirmed that the a_w decreasing effect of gelatin is less harmful to cells than the effect of Na⁺ ions. The unexpected shift of the growth/no growth interface to more severe conditions when going from a liquid medium to a medium with 10 g L⁻¹ gelatin is more pronounced when formulating the models in terms of a_w than in terms of NaCl concentrations. At 50 g L⁻¹ gelatin, the model factored with NaCl concentration shifts to milder conditions (concordant to literature results) while the model with a_w indicates a further shift to more severe conditions, which is due to the water activity lowering effect of gelatin and the interaction between gelatin and NaCl. The results suggest that solute concentration should be used instead of a_w, both for kinetic models in the growth region and for growth/no growth interface models, if the transferability of models to solid foods is to be increased.     

Development of a novel method for feeding a mixture of -lactic acid and acetic acid in fed-batch culture of Ralstonia eutropha for poly- -3-hydroxybutyrate production

Fermentative production of poly- -3-hydroxybutyrate [P(3HB)] from a mixture of -lactic acid and acetic acid by Ralstonia eutropha was investigated. For fed-batch culture with cell density, it is necessary to control the concentration of these organic acids in the culture medium below the inhibitory level for cell growth. Therefore, a novel feeding method, termed the computer-controlled pH-stat substrate feeding method, was developed using the rate of increase of the pH (pH-increasing rate) of the culture medium as an indicator for feed control. The pH-increasing rate, which was calculated every minute by a pH meter-linked computer, represented secondary information regarding substrate consumption by cells. When the pH-increasing rate decreased to 5% of the maximum increasing rate, acidic substrate solution was fed into the fermentor until the pH was reduced to 7.00. Using this feeding strategy, the cell concentration and PHA content obtained in 42 h were 75.0 g/l and 73.1% (w/w), respectively, resulting in a high P(3HB) productivity of 1.30 g/l·h.     

Triglycerides constituted of short and medium chain fatty acids and dicarboxylic acids in Momordica charantia, as well as capric acid,inhibit PGE2 production in RAW264.7 macrophages

This study was aimed to identify compounds in bitter gourd (Momordica charantia) ethyl acetate extract (EAE), which inhibit lipopolysaccharide-induced prostaglandin E₂ (PGE₂) production in RAW264.7 cells. Bitter gourd EAE was partitioned between n-hexane and methanol/H₂O (90/10). The hexane fraction was further separated by repeated silica gel chromatographies, and a reverse phase (RP) C18 chromatography. Fraction RP-10 showed the highest inhibition effect on PGE₂ production (Max inhibition = 96%, IC₅₀ = 2.3 μg/ml) and was identified to be triglycerides constituted of short and medium chain fatty acids by ¹H NMR, IR and H–HCOSY, and dicarboxylic acids by GC/MS. Fatty acids with 3–20 carbons were tested for the inhibitory activity, and capric acid exhibited the highest effect (Max inhibition = 99%, IC₅₀ = 6.5 μM). In conclusion, triglycerides composed of short and medium chain fatty acids and dicarboxylic acids in bitter gourd inhibit PGE₂ production, and capric acid is the most potent inhibitor among the fatty acids.     

Fermentation of rice using amylolytic Bifidobacterium

Twenty-two amylolytic bifidobacteria grown in BHI-starch media were compared for the amylase activity of the intra- and extra-cellular enzymes. The activity of the cells grown in the liquid medium differed considerably. Among the strains tested B. adolescentis Int57 and B. adolescentis ZS8 exhibited higher activities than others. In rice medium containing 0.05% -cysteine·HCl and 0.2% yeast extract, the amylolytic Bifidobacterium strains grew considerably better than non-amylolytic Bifidobacterium strains. B. adolescentis Int57, which showed highest growth and amylase activity in the rice medium, was chosen and rice fermentation was investigated. The titratable acidity (TA) reached 2.43 and pH decreased to 4.4 after 24 h fermentation. The relative ratio of acetic acid to lactic acid gradually decreased during fermentation. The concentration of reducing sugar and amylase activity gradually increased and reached 14 mg maltose equivalent/ml and 35 mU/ml min, respectively, in 24 h. The accumulated reducing sugar was mostly maltotriose. The layer-separation of fermented product was stabilized by the addition of 1% gelatin. It was suggested that amylolytic bifidobacteria may be used for the production of fermented rice products.     

Initial screening for inhibitory activity of essential oils on growth of Fusarium verticillioides, F. proliferatum and F. graminearum on maize-based agar media

An in vitro initial screening of a range of 37 essential oils on inhibition of mycelial growth of Fusarium verticillioides, F. proliferatum and F. graminearum under different temperature (20–30°C) and water activity (a_w) (0.95–0.995) conditions was made. The basic medium was a 3% maize meal extract agar. The maize meal agar was modified with glycerol to a range of water activity conditions and the essential oils were incorporated at different concentrations (0, 500, 1000 μg ml⁻¹). Cinnamon leaf, clove, lemongrass, oregano and palmarosa oils were the products tested suitable for being used as novel preservatives in the control of the three Fusarium species studied. Although water activity was determinant for the growth of the isolates, in general, the preservative effects of the oils were not linked to a_w. However, a trend to a higher inhibition by the oils when a_w was low was observed. Temperature had a minor importance in the inhibitory effect of the preservatives. In vivo studies may be required to confirm the usefulness of the results obtained.     

Effect of growing bacteria isolated from food on staphylococcal growth and thermonuclease activity

The effect of growing Bacillus subtilis, Streptococcus faecalis var. liquefaciens, Escherichia coli and Lactobacillus plantarum on staphylococcal growth and thermonuclease (TNase) activity was investigated in liquid media and in foods. Growth of S. aureus at 37°C for 24 h under aerobic conditions was not inhibited by the four test strains. However, staphylococcal TNase activity decreased by 70 and 80% in the presence of B. subtilis and S. faecalis var. liquefaciens respectively. Staphylococcal growth and TNase activity were strongly inhibited by L. plantarum under anaerobic conditions at pH 5.5 but not at pH 7.0. Furthermore, optimal TNase production by S. aureus occurred in cooked meat medium containing 0.5 to 5.0% NaCl. TNase production significantly decreased at higher concentrations of NaCl. In the presence of B. subtilis and S. faecalis var. liquefaciens. TNase activity decreased at NaCl levels of 0.5 to 5.0% but not at NaCl concentrations>5.0%. TNase activity was also inhibited by growing B. subtilis and S. faecalis var. Liquefaciens at pH 5.0 to 7.0. The rate of inhibition increased with increasing pH. TNase activity was not inhibited after 48 h incubation at 20° in the presence of B. subtilis and S. faecalis var. liquefaciens but significant inactivation could be demonstrated at 25° to 37°C. The results obtained with artificially contaminated, sterile food samples were similar to those obtained with brain-heart infusion broth, but the degree of decrease in TNase activity in food was much lower than that in brain-heart infusion broth.     

The effect of nisin and garlic (Allium sativum L.) essential oil separately and in combination on the growth of Listeria monocytogenes

In the present study, the anti-listerial activity of the nisin and garlic (Allium sativum L.) essential oils (GO) alone and in combination was investigated at different temperatures (20 and 30 °C), pH (6.8, 5.6 and 4.8) and NaCl concentrations (0, 0.5, 2.5 and 4.5 g/100 mL). Minimum inhibitory concentrations (MICs) against Listeria monocytogenes were assessed for the nisin and essential oil. Furthermore, for combinations of the antimicrobials, the Differences in Population (DP) method were used to determine their effect. Both essential oil and nisin possessed considerable antimicrobial effects on the microorganism. The MICs for nisin and GO were 12.5 IU/mL and 100 μg/mL, respectively. Regardless of NaCl concentration and temperature, the anti-listerial activity of both GO and nisin was strongly influenced by pH. Moreover, at the same temperature, and regardless of pH value, the growth of the organism was also affected by increasing NaCl concentration. In combination, the DP was related strongly to the agent’s concentration and media pH. Meanwhile, among all combined systems, the effect of the combination (EC) of nisin with GO at 30 °C, pH 5.6 and 0 g/100 mL NaCl, showed significant anti-listerial activity (P ≤ 0.05).     

Antagonistic effect of fat on the antibotulinal activity of food preservatives and fatty acids

The effect of fat on the antibotulinal activity of 11 food preservatives, 12 free fatty acids, and nine lots of enzyme-modified cheese (EMC) was evaluated in a media system. Anhydrous milkfat or soybean oil was added to tubes of Trypticase–peptone–glucose–yeast extract medium (TPGY) supplemented with the additives (final pH adjusted to 5.9). Treatments were inoculated with 3-log₁₀ proteolytic Clostridium botulinum spores/ml (10-strain mixture of serotypes A and B) and incubated anaerobically at 30°C for up to 14 days. For the preservative and fatty acids studies, growth of C. botulinum was determined by measuring optical changes at OD_{640 nm}. Botulinal toxin production was determined in EMC-treatments using the mouse bioassay. Data revealed that the antibotulinal effects of nisin, and free fatty acids caprylic, capric, lauric, myristic, oleic, and linoleic acids were each significantly reduced in treatments supplemented with 20% fat (P<0.05). Similar trends were observed in TPGY supplemented with 20% fat and potassium sorbate, sorbic acid, monolaurin, polyphosphate emulsifier, or EDTA–lysozyme, but the differences were reduced. Fat was also antagonistic to the antibotulinal activity of five EMC-treatments. This study suggests that fat may reduce the efficacy of some antimicrobials added to or found naturally in foods.     

Inhibitory effect of Cynara cardunculus L. extract on aflatoxin B1 production by Aspergillus parasiticus in sesame (Sesamum indicum L.)

Aflatoxin B₁ has been considered as the most potent liver carcinogen for humans. One factor that stimulates the production of aflatoxins in oilseed products, such as sesame seeds and their products is the presence of high levels of fatty acids. This work presents further evidence that sesame is a favorable substrate for aflatoxin B₁ production by Aspergillus parasiticus. Moreover, the use of wild artichoke (Cynara cardunculus L.) extract was examined showing inhibition of aflatoxin B₁ production in both sesame and yeast extract sucrose medium, inoculated with A. parasiticus. More specifically, the inhibition of aflatoxin B₁ production by A. parasiticus in sesame seeds paste was 99.6% and in yeast extract sucrose medium it was 99.4%.     

Inactivation of Yersinia enterocolitica by pulsed electric fields

The influence of growth conditions, treatment medium characteristics and PEF process parameters on the lethal effect on Yersinia enterocolitica of pulsed electric fields (PEF) treatments in batch has been investigated. Growth phase, temperature of growth, pH, conductivity of the treatment medium, pulse width and frequency of pulses did not influence the sensitivity of Y. enterocolitica to PEF. However, an A_w decrease from >0.99 to 0.93 of the treatment medium increased the PEF resistance of Y. enterocolitica with 3.5 log₁₀ cycles after a treatment of 22 kV/cm, 800 μs and 880 kJ/kg. Inactivation of Y. enterocolitica increased with the field strength, treatment time and total specific energy up to a maximum of 6 log₁₀ cycles after 28 kV/cm, 2000 μs and 3559 kJ/kg. A nonlinear relationship was found among the survival fraction and the treatment time or the specific energy that was accurately described by a mathematical model based on the Weibull distribution. The inactivation of Y. enterocolitica by PEF was characterized by maximum field strength thresholds. Above these thresholds, specific energy necessary to obtain a given level of inactivation scarcely decreased by increasing the electric field strength, and inactivation of Y. enterocolitica only depended on the specific energy applied.     

Extractability of African yam bean (Sphenostylis stenocarpa) protein in acid, salt and alkaline aqueous media

Protein extractability from defatted Africa yam been (Sphenostylis stenocarpa) was studied under various conditions: solid/solvent ratio, time, pH, and salt. Extractable protein from S. stenocarpa was strongly dependent on all these factors. Maximum extractable protein was obtained after 2 h extraction time; the solid to solvent ratio in the range of 1:20–1:50 gave maximum protein extractability. The pH corresponding to maximum and minimum extractable proteins were 10 and 5, respectively, but addition of NaCl changed this slightly. Extractable protein of 92%, 88% and 84% were obtained in aqueous concentration of 0.01 MNa₂SO₄, 1.5 MNaCl and 0.01 MNaOH, respectively while other concentrations gave lower extractability. These salt concentrations, i.e. 0.01 MNa₂SO₄ and 1.5 MNaCl gave slightly lower extractable protein under alkaline condition. S. stenocarpa flour has a higher buffer capacity in acid medium than in alkaline medium.     

Improvement of exo-polysaccharides production and modeling kinetics by Armillaria luteo-virens Sacc. in submerged cultivation

Armillaria luteo-virens Sacc. is a special medicinal and edible mushroom in China, and its wild type has been investigated that it contains some functional components. The optimization of submerged culture conditions for mycelium growth and exo-polysaccharide (EPS) production in an edible mushroom A. luteo-virens Sacc. was studied in shake flasks and bioreactors with response surface methodology (RSM). A fractional factorial design (FFD) and central composite design (CCD) were applied to optimize the main factors that affect the fungal polysaccharides production in a submerged culture. Glucose is very important for EPS production by A. luteo-virens Sacc., and yeast extract powder also significantly affects fungal EPS production through fractional factorial experiment. The optimized cultivation medium constituents were derived from the predicted model using central composite design as follows: 31.26 g/l glucose, 1.06 g/l yeast extract powder, 1.00 g/l K₂HPO₄, 0.50 g/l KH₂PO₄, and 0.45 g/l MgSO₄·7H₂O, pH unadjusted. The predicted maximal EPS concentration was 5.40 g/l using the optimized culture medium compositions at 23 °C for 5 days. Additionally, the submerged cultivation process was explained using the optimized cultivation medium. The cultivation kinetics model constructed was suitably depicting the culture process in bioreactor cultivation scale.     

吡哆醇对鲁氏接合酵母高盐胁迫耐受的影响

为了提高鲁氏接合酵母抗高盐胁迫能力,作者探索了添加吡哆醇对鲁氏接合酵母高盐胁迫耐受的影响。通过测量甘油、海藻糖含量以及Na⁺/K⁺-ATP酶活性等一系列生理表征来验证鲁氏接合酵母高盐适应性的变化。实验结果表明,鲁氏接合酵母在高盐胁迫条件下外源添加吡哆醇,在延滞期时,菌株提前24 h积累细胞内甘油,并且加快了细胞内海藻糖分解代谢速率;在对数期时,细胞内钠钾比率（Na⁺/K⁺）降低82.3%,细胞膜上Na⁺/K⁺-ATP酶活性增加16.9%,2-苯乙醇产量（质量浓度）提高6.42倍;在稳定期时,生物量提高10.6%,乙醇产量（质量浓度）提高5%,2-苯乙醇产量（质量浓度）提高1.26倍。综上所述,吡哆醇的添加能有效提高鲁氏接合酵母的高盐胁迫耐受能力,进一步增强了鲁氏接合酵母在酱油等高盐发酵食品中的应用前景。